Visualized Detection of Vibrio parahaemolyticus in Food Samples Using Dual-Functional Aptamers and Cut-Assisted Rolling Circle Amplification

摘要

A biosensor using two aptamers (Dual-Apt) and cut-assisted rolling circle amplification (CA-RCA) for rapid and visualized detection of Vibrio parahaemolyticus was established. The anchoring aptamer (A-Apt) that specifically binds to the surface of V. parahaemolyticus was applied to separate and enrich the bacterium from the food matrix with the help of streptavidin magnetic beads. While the detecting aptamer (D-Apt), binding on the different sites of the cell surface, was used as a signal reporter. CA-RCA with an enhanced amplification rate was fabricated here to amplify the D-Apt to produce the monomeric G4 sequence that catalyzes the oxidation of ABTS(2)(-), resulting in the coloration visible to the naked eye. Under optimal conditions, as low as 10 colony-forming units (CFU)/mL (g) of V. parahaemolyticus can be visibly detected in real food samples. Free from DNA extraction, visualized signal output and no need for expensive instruments enable Dual-Apt and CARCA to be a promising strategy for on-spot rapid detection.